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Recombinant human granulocyte colony-stimulating factor augments cytotoxicity of OK-432-induced polymorphonuclear
1First Department of Internal Medicine, Kyoto Prefectural University of Medicine, Japan.
Abstract:
Polymorphonuclear leukocytes (PMNs) recovered from the peritoneal cavity of mice treated with the streptococcal preparation OK-432, exhibited strong cytotoxicity after the in vitro addition of Nocardia rubra cell wall skeleton (N-CWS). In this study, we investigated whether recombinant human granulocyte colony-stimulating factor (rhG-CSF) could augment the cytotoxicity of OK-432-induced PMNs after the addition of N-CWS in vitro. PMNs recovered from the peritoneal cavity of 8- to 10-week-old, male C3H/He mice induced by intraperitoneal (i.p.) injection of 50 KE/kg (1 KE = 0.1 mg) of OK-432 were used in a 51Cr release assay against MM46 mammary carcinoma cells. While addition of rhG-CSF in vitro did not augment the cytotoxicity of OK-432-induced PMNs, marked augmentation of the cytotoxicity of OK-432-induced PMNs was observed following a single subcutaneous (s.c.) or i.p. injection of 125 micrograms/kg of rhG-CSF. The effect of in vivo administered rhG-CSF was dependent on the timing of the injection with respect to OK-432 administration and differed from s.c. or i.p. injections. Interestingly, the cytotoxicity of OK-432-induced PMNs was rather weak following consecutive s.c. or i.p. administration of rhG-CSF for 7-14 days. H2O2 is likely involved in mediating the cytotoxicity of OK-432-induced PMNs since activity was significantly reduced by the in vitro addition of low concentration of catalase. Generation of H2O2 by the PMNs correlated with cytotoxicity. These results suggest that in vivo administration of rhG-CSF augments the cytotoxicity of OK-432-induced PMNs in a time dependent fashion and that H2O2 plays an important role in mediating their cytotoxicity.
Insights
Recombinant human granulocyte colony-stimulating factor (rhG-CSF) enhances the cancer-fighting ability of immune cells when injected into mice, but not when added in vitro. This augmentation is time-dependent and involves hydrogen peroxide (H2O2) production.
Area of Science:
- Immunology
- Cancer Research
- Pharmacology
Background:
- Streptococcal preparation OK-432 induces polymorphonuclear leukocytes (PMNs) with cytotoxic activity.
- Nocardia rubra cell wall skeleton (N-CWS) enhances PMN cytotoxicity in vitro.
- Recombinant human granulocyte colony-stimulating factor (rhG-CSF) is a growth factor for neutrophils.
Purpose of the Study:
- To investigate if rhG-CSF can augment the cytotoxicity of OK-432-induced PMNs against cancer cells.
- To determine the optimal administration route and timing for rhG-CSF to enhance PMN-mediated cytotoxicity.
- To explore the role of hydrogen peroxide (H2O2) in rhG-CSF-mediated augmentation of PMN cytotoxicity.
Main Methods:
- PMNs were isolated from mice treated with OK-432.
- Cytotoxicity was assessed using a 51Cr release assay against MM46 mammary carcinoma cells.
- rhG-CSF was administered in vitro, or via subcutaneous (s.c.) or intraperitoneal (i.p.) injection at various times relative to OK-432 treatment.
- The role of H2O2 was investigated using catalase.
Main Results:
- In vitro addition of rhG-CSF did not enhance PMN cytotoxicity.
- Single s.c. or i.p. injections of rhG-CSF significantly augmented PMN cytotoxicity.
- The timing of rhG-CSF administration was crucial for its augmenting effect.
- Consecutive rhG-CSF administration for 7-14 days resulted in weak cytotoxicity.
- H2O2 generation by PMNs correlated with cytotoxicity and was essential for the effect.
Conclusions:
- In vivo administration of rhG-CSF augments the cytotoxicity of OK-432-induced PMNs in a time-dependent manner.
- The route and timing of rhG-CSF administration significantly influence its efficacy.
- Hydrogen peroxide plays a critical role in mediating the enhanced cytotoxicity.